Hearts of Oak joined the list of second round casualties in the MTN FA Cup as they became the second Premier League team to have been eliminated.
And they join a list of five elite casualties so far, coming a day after Wa All Stars were eliminated from the competition.
Other Premier League teams who have been dumped out of the competition by teams from the lower divisions are Ebusua Dwarfs, Heart of Lions and Amidaus Professionals.
The Phobians were kicked out on Sunday after a 2-1 defeat to Division One league side, Gold Stars in Bibiani.
It will come as a blow to their title ambitions as their elimination leaves Mohammed Polo with only one silverware - Premier League title - to compete for now.
The defeat also represents their second-straight loss in a competitive match following their 3-0 loss to Wa All Stars seven days ago in the Premier League.
However, unlike the Phobians, reigning Cup holders Medeama, AshantiGold and Berekum Chelsea survived testing battles to progress to the third round.
Medeama defeated Division Two side, Score FC 2-1 in Koforidua while AshantiGold were 3-1 winners against former premiership side, B.A. United.
At the Golden City, Berekum Chelsea won 3-0 over Wenchi United.
Round of 64 Results
Matches played on December 22
Berekum Chelsea 3-0 Wenchi United
Gold Stars 2-1 Accra Hearts of Oak
AshantiGold 3-1 BA United
Score FC 1-2 Medeama
Dolphins FC 2-2(4-2 pen) Guan United
Prestige FC 0-1 Istanbul FC
New Edubiase 1-0 Bolga Man City
Tarkwa United 2-0 Hearts of Lions
Ebusua Dwarfs 1-1(4-5 pen) First Klass FC
Okwawu United 1-0 Real Athletico
King Faisal 3-1 Tuobodom Santos
Starke FC 1-2 Hasaacas
Pure Joy 2-1 Ajax Fc
Bechem United 4-0 Cape Coast Strikers
Amidaus Professionals 0-2 Danbort FC